Oxygen toxicity in control and H2O2-resistant Chinese hamster fibroblast cell lines

Spitz, Douglas R.; Elwell, James H.; Sun, Yi; Oberley, Larry W.; Oberley, Terry D.; Sullivan, Shannon J.; Roberts, Robert J.

doi:10.1016/0003-9861(90)90489-l

Cited by 99 publications

(81 citation statements)

References 33 publications

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“…Primary antibodies raised against bovine liver catalase, CuZnSOD, human kidney MnSOD, and GPX were characterized; their specificities were determined previously. 32 Immunoblot analyses of proteins from frozen liver tissues were performed after homogenization of small aliquots of liver tissue in ice-cold hypotonic buffer containing 10 mmol/L Tris-HCl (pH 7.6), 2 mmol/L ethylenediaminetetraacetic acid, and 4 mmol/L MgCl 2 in the presence of a protease-inhibitor cocktail (10 µg/mL each of aprotinin, pepstatin, leupeptin, and 5 µg/mL of phenylmethylsulfonylfluoride). The resulting supernatant was subjected to differential centrifugation at 500g for 20 minutes to isolate cytosolic and crude membrane preparations and nuclear fractions.…”

Section: Methodsmentioning

confidence: 99%

Increased lipid peroxidation and impaired antioxidant enzyme function is associated with pathological liver injury in experimental alcoholic liver disease in rats fed diets high in corn oil and fish oil

et al. 1998

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Increased hepatic oxidative stress with ethanol administration is hypothesized to be caused either by enhanced pro-oxidant production or decreased levels of antioxidants or both. We used the intragastric feeding rat model to assess the relationship between hepatic antioxidant enzymes and pathological liver injury in animals fed different dietary fats. Male Wistar rats (5 per group) were fed ethanol with either medium-chain triglycerides (MCTE), palm oil (PE), corn oil (CE), or fish oil (FE). Control animals were fed isocaloric amounts of dextrose instead of ethanol with the same diets. The following were evaluated in each group: liver pathology, lipid peroxidation, manganese superoxide dismutase (MnSOD) levels, copper-zinc SOD (CuZnSOD) levels, glutathione peroxidase (GPX) levels, and catalase (CAT) levels. All enzymes were evaluated using activity assays and immunoblots. Rats fed FE showed the most severe pathology (fatty liver, necrosis, and inflammation), those fed CE showed moderate changes, those fed PE showed fatty liver only, and those fed MCTE were normal. Parameters indicative of lipid peroxidation (conjugated dienes and thiobarbituric acid-reactive substances) were also greater in rat livers from animals fed the diets high in polyunsaturated fatty acids (CE and FE). CuZnSOD, GPX, and CAT activities showed an inverse correlation (r ‫؍‬ ؊.92, P F .01) with severity of pathological injury, with the lowest levels for both enzymes found in FE-fed rats. Decreased enzyme activity in CE-and FE-fed rats was accompanied by similar decreases in immunoreactive protein. Ethanol administration did not cause significant decreases in enzyme activity in groups that showed no necroinflammatory changes (MCTE and PE). MnSOD activity showed no significant change in any ethanol-fed group. Our results show that decreases in CuZnSOD, GPX, and CAT occur in rats showing pathological liver injury and also having the highest levels of lipid peroxidation. These results suggest that feeding dietary substrates that enhance lipid peroxidation can exacerbate both ethanol-induced oxidative damage as well as necroinflammatory changes. The decrease in activity of antioxidant enzymes observed in animals fed diets high in polyunsaturated fatty acids and ethanol could possibly increase the susceptibility to oxidative damage and further contribute to ethanol-induced liver injury. (HEPATOLOGY 1998;27:1317-1323.)

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Section: Methodsmentioning

confidence: 99%

Increased lipid peroxidation and impaired antioxidant enzyme function is associated with pathological liver injury in experimental alcoholic liver disease in rats fed diets high in corn oil and fish oil

et al. 1998

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“…Catalase activity was determined on whole homogenates by measuring the disappearance of 10 mM hydrogen peroxide (Δε 240 = 39.4 M −1 cm −1 ) in 50 mM potassium phosphate, pH 7.0, monitored at 240 nm and the units were expressed as milli-k units/mg of protein as described [31].…”

Section: Catalase Activity Assaymentioning

confidence: 99%

2-Deoxyglucose combined with wild-type p53 overexpression enhances cytotoxicity in human prostate cancer cells via oxidative stress

Ahmad

Abdalla

Aykin-Burns

et al. 2008

Free Radical Biology and Medicine

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Over expression of the tumor suppressor gene, wild type p53 (wtp53), using adenoviral vectors (Adp53) has been suggested to kill cancer cells by hydroperoxide-mediated oxidative stress [1,2] and nutrient distress induced by the glucose analog, 2-deoxyglucose (2DG), has been suggested to enhance tumor cell killing by agents that induce oxidative stress via disrupting hydroperoxide metabolism [3,4]. In the current study clonogenic cell killing of PC-3 and DU-145 human prostate cancer cells (lacking functional p53) mediated by 4 h exposure to 50 plaque forming units (pfus)/ cell of Adp53 (that caused the enforced over expression of wtp53) was significantly enhanced by treatment with 2DG. Accumulation of glutathione disulfide was found to be significantly greater in both cell lines treated with 2DG+Adp53 and both cell lines treated with 2DG+Adp53 showed a ~2-fold increases in dihydroethidine (DHE) and 5-(and-6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate (CDCFH 2 ) oxidation, indicative of increased steady-state levels of O 2 •-and hydroperoxides, respectively. Finally, over expression of catalase or glutathione peroxidase using adenoviral vectors partially, but significantly, protected DU-145 cells from the toxicity induced by 2DG+Adp53 treatment. These results show that treatment of human prostate cancer cells with the combination of 2DG (a nutrient stress) and over expression of the tumor suppressor gene, wtp53, enhances clonogenic cell killing by a mechanism that involves oxidative stress as well as allowing for the speculation that inhibitors of glucose and hydroperoxide metabolism can be used in combination with Adp53 gene therapy to enhance therapeutic responses.

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“…32 Total superoxide dismutase (SOD) activity of liver homogenates was measured by a modification of the nitro blue tetrazolium method, as described previously. 32,33 One unit of SOD activity was defined as the amount of protein required to inhibit the reduction of nitro blue tetrazolium by 50%. 32 …”

Section: Other Antioxidant Enzyme Assaysmentioning

confidence: 99%

“…32,33 One unit of SOD activity was defined as the amount of protein required to inhibit the reduction of nitro blue tetrazolium by 50%. 32 …”

Section: Other Antioxidant Enzyme Assaysmentioning

confidence: 99%

Deficiency of Glutathione Peroxidase-1 Sensitizes Hyperhomocysteinemic Mice to Endothelial Dysfunction

Dayal

Brown

Weydert

et al. 2002

ATVB

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100

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Objective-We tested the hypothesis that deficiency of cellular glutathione peroxidase (GPx-1) enhances susceptibility to endothelial dysfunction in mice with moderate hyperhomocysteinemia. Methods and Results-Mice that were wild type (Gpx1 ϩ/ϩ ), heterozygous (Gpx1 ϩ/Ϫ ), or homozygous (Gpx1 Ϫ/Ϫ ) for the mutated Gpx1 allele were fed a control diet or a high-methionine diet for 17 weeks. Plasma total homocysteine was elevated in mice on the high-methionine diet compared with mice on the control diet (23Ϯ3 versus 6Ϯ0.3 mol/L, respectively; PϽ0.001) and was not influenced by Gpx1 genotype. In mice fed the control diet, maximal relaxation of the aorta in response to the endothelium-dependent dilator acetylcholine (10 Ϫ5 mol/L) was similar in Gpx1 ϩ/ϩ , Gpx1 ϩ/Ϫ , and Gpx1 Ϫ/Ϫ mice, but relaxation to lower concentrations of acetylcholine was selectively impaired in Gpx1 Ϫ/Ϫ mice (PϽ0.05 versus Gpx1 ϩ/ϩ mice). In mice fed the high-methionine diet, relaxation to low and high concentrations of acetylcholine was impaired in Gpx1 PϽ0.05). No differences in vasorelaxation to nitroprusside or papaverine were observed between Gpx1 ϩ/ϩ and Gpx1 Ϫ/Ϫ mice fed either diet. Dihydroethidium fluorescence, a marker of superoxide, was elevated in Gpx1 Ϫ/Ϫ mice fed the high-methionine diet (PϽ0.05 versus Gpx1 ϩ/ϩ mice fed the control diet). Conclusions-These findings demonstrate that deficiency of GPx-1 exacerbates endothelial dysfunction in hyperhomocysteinemic mice and provide support for the hypothesis that hyperhomocysteinemia contributes to endothelial dysfunction through a peroxide-dependent oxidative mechanism. Key Words: endothelium Ⅲ homocysteine Ⅲ nitric oxide Ⅲ peroxide H yperhomocysteinemia is an emerging risk factor for cardiovascular events and venous thrombosis, 1,2 but the mechanisms responsible for the vascular pathology of hyperhomocysteinemia are still incompletely understood. Like many other cardiovascular risk factors, hyperhomocysteinemia produces endothelial dysfunction, which is possibly due to oxidative inactivation of endothelium-derived NO. 3,4 The oxidative stress hypothesis 4 is supported by the observation that auto-oxidation of homocysteine in vitro generates reactive oxygen species (ROS), including hydrogen peroxide and superoxide, and promotes oxidation of LDL. 5-7 Treatment of cultured endothelial cells with homocysteine decreases bioavailable NO 8,9 and produces cytotoxicity mediated by hydrogen peroxide. 10 Homocysteine also indirectly contributes to oxidative stress by inhibiting the expression of antioxidant enzymes, such as cellular glutathione peroxidase (GPx-1), an effect that may sensitize one to the toxic effects of homocysteine-derived hydrogen peroxides and lipid peroxides. 8,11 However, the role of oxidative stress in the vascular dysfunction of hyperhomocysteinemia in vivo is less clear, and its clinical importance has been questioned. 12 Attempts to demonstrate elevated levels of oxidation products in humans with hyperhomocysteinemia have produced conflicting results. [13][14][15][16] R...

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Oxygen toxicity in control and H2O2-resistant Chinese hamster fibroblast cell lines

Cited by 99 publications

References 33 publications

Increased lipid peroxidation and impaired antioxidant enzyme function is associated with pathological liver injury in experimental alcoholic liver disease in rats fed diets high in corn oil and fish oil

Increased lipid peroxidation and impaired antioxidant enzyme function is associated with pathological liver injury in experimental alcoholic liver disease in rats fed diets high in corn oil and fish oil

2-Deoxyglucose combined with wild-type p53 overexpression enhances cytotoxicity in human prostate cancer cells via oxidative stress

Deficiency of Glutathione Peroxidase-1 Sensitizes Hyperhomocysteinemic Mice to Endothelial Dysfunction

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